Substituent activity relationship studies on new azolo benzoxazepinyl oxazolidinones.

In an effort to discover potent antibacterials based on the entropically favored 'bioactive conformation' approach, a series of novel tricyclic molecules mimicking the conformationally constrained structure of Linezolid is reported. Based on the initial tricyclic molecule 1, the benzazepine derivative 2 was designed where the tricyclic structure had more flexibility around C-N bond compared to 1. While, the molecule 2 was less active, the molecule 3 showed promising antibacterial activity presumably after having obtained rigidity due to pyrrole ring. The syntheses, SAR studies, and evaluation of 3 as a lead compound are reported.

Effect of ultraviolet B (302 nm) irradiation on viability, metabolic and detoxification functions of goat hepatocytes--in vitro study.

The object of the present study was to investigate the effect(s) of UV-B irradiation on the functional integrity, metabolic and detoxifying capacity of the isolated goat hepatocytes. Isolated goat hepatocytes were subjected to UV-B irradiation invitro for 0, 250, 500, 1250, 2500 and 7500 Joules/m2 which correspond to the irradiation time of 0, 1, 2, 5, 10 and 30 min. Cells were then analysed for Viability (Trypan blue exclusion test [TBE], 3-[4,5-dimethylthiozol-2yl]-2,5-diphenyltetrazolium bromide [MTT] assay, Membrane integrity (Lactate dehydrogenase [LDH] leakage, Lipid peroxidation) Detoxification (Ureagenesis, Cytochrome P450 activity [CYP450, Diazepam metabolism] and Glutathione-S-Transferase [GST] activity. The results show that there was no difference in functional, metabolic as well as detoxifying parameters of the hepatocytes when irradiated from 0-1250 Joules/m2, whereas a significant alteration was appreciable in the parameters such as LDH leakage, lipid peroxidation, and CYP450 activity when irradiated beyond 1250 Joules/m2. Our present findings suggest that the biologically compatible and feasible dose of UV-B irradiation for xenotransplantation appears to be 1250 Joules/m2.

Intravenous pharmacokinetics, oral bioavailability and dose proportionality of ragaglitazar, a novel PPAR-dual activator in rats.

Pharmacokinetics of ragaglitazar (a novel phenoxazine derivative of aryl propanoic acid), a potent insulin sensitizing and lipid-lowering compound was studied in Wistar rats. A single dose of 1, 3 or 10 mg/kg of ragaglitazar was given orally to male rats (n=4 per dose level) to evaluate dose proportionality. In another study, a single intravenous bolus dose of ragaglitazar was given to rats (n=4) at 3 mg/kg dose following administration through the lateral tail vein in order to obtain the absolute oral bioavailability and clearance parameters. Blood samples were drawn at predetermined intervals and the concentration of ragaglitazar in plasma was determined by a validated HPLC method. Plasma concentration versus time data were generated following oral and intravenous dosing and pharmacokinetic analysis was performed using non-compartmental analysis. The results revealed that Cmax and AUC(0-infinity) increased more than proportionally to the administered oral doses. As dose increased in the ratio of 1:3:10, the mean Cmax and AUC(0-infinity) increased in the ratio of 1:3.2:13 and 1:3.2:16, respectively. After intravenous administration the systemic clearance and volume of distribution of ragaglitazar in rats were 139+/-30 ml/h/kg and 463+/-51 ml/kg, respectively (mean+/-SD). Plasma concentrations declined mono-exponentially following intravenous administration and elimination half-life (t1/2) was about 2.6 h and not significantly different (p > 0.05) from the value from oral administration. Mean residence time (MRT) values for ragaglitazar were found to be 4.15+/-0.52 h (3.5 to 4.6 h). Absolute oral bioavailability of ragaglitazar across the doses tested was in the range of 68%-93%. In conclusion, ragaglitazar exhibits promising pharmacokinetic properties in rats.

Development and validation of a chiral liquid chromatographic method, based on Chiralpak to quantify enantiomers of (+/-)-DRF 2725 in rat plasma: lack of inversion of ragaglitazar (S(-)-DRF 2725) to its antipode in plasma.

A selective, accurate and reproducible high-performance liquid chromatographic (HPLC) method for the separation of individual enantiomers of DRF 2725 [R(+)-DRF 2725 and S(-)-DRF 2725 or ragaglitazar] was obtained on a chiral HPLC column (Chiralpak). During method optimization, the separation of enantiomers of DRF 2725 was investigated to determine whether mobile phase composition, flow-rate and column temperature could be varied to yield the base line separation of the enantiomers. Following liquid-liquid extraction, separation of enantiomers of DRF 2725 and internal standard (I.S., desmethyl diazepam) was achieved using an amylose based chiral column (Chiralpak AD) with the mobile phase, n-hexane-propanol-ethanol-trifluoro acetic acid (TFA) in the ratio of 89.5:4:6:0.5 (v/v). Baseline separation of DRF 2725 enantiomers and I.S., free from endogenous interferences, was achieved in less than 25 min. The eluate was monitored using an UV detector set at 240 nm. Ratio of peak area of each enantiomer to I.S. was used for quantification of plasma samples. Nominal retention times of R(+)-DRF 2725, S(-)-DRF 2725 and I.S. were 15.8, 17.7 and 22.4 min, respectively. The standard curves for DRF 2725 enantiomers were linear (R(2) > 0.999) in the concentration range 0.3-50 microg/ml for each enantiomer. Absolute recovery, when compared to neat standards, was 70-85% for DRF 2725 enantiomers and 96% for I.S. from rat plasma. The lower limit of quantification (LLOQ) for each enantiomers of DRF 2725 was 0.3 microg/ml. The inter-day precisions were in the range of 1.71-4.60% and 3.77-5.91% for R(+)-DRF 2725, S(-)-DRF 2725, respectively. The intra-day precisions were in the range of 1.06-11.5% and 0.58-12.7% for R(+)-DRF 2725, S(-)-DRF 2725, respectively. Accuracy in the measurement of quality control (QC) samples was in the range 83.4-113% and 83.3-113% for R(+)-DRF 2725, S(-)-DRF 2725, respectively. Both enantiomers and I.S. were stable in the battery of stability studies viz., bench-top (up to 6 h), auto-sampler (up to 12 h) and freeze/thaw cycles (n = 3). Stability of DRF 2725 enantiomers was established for 15 days at -20 degrees C. The application of the assay to a pharmacokinetic study of ragaglitazar [S(-)-DRF 2725] in rats is described. It was unequivocally demonstrated that ragaglitazar does not undergo chiral inversion to its antipode in vivo in rat plasma.

Synthesis and cyclooxygenase-2 inhibiting property of 1,5-diarylpyrazoles with substituted benzenesulfonamide moiety as pharmacophore: Preparation of sodium salt for injectable formulation.

A series of 1,5-diarylpyrazoles having a substituted benzenesulfonamide moiety as pharmacophore was synthesized and evaluated for cyclooxygenase (COX-1/COX-2) inhibitory activities. Through SAR and molecular modeling, it was found that fluorine substitution on the benzenesulfonamide moiety along with an electron-donating group at the 4-position of the 5-aryl ring yielded selectivity as well as potency for COX-2 inhibition in vitro. Among such compounds 3-fluoro-4-[5-(4-methoxyphenyl)-3-trifluoromethyl-1H-1-pyrazolyl]-1-benzenesulfonamide 3 displayed interesting pharmacokinetic properties along with antiinflammatory activity in vivo. Among the sodium salts tested in vivo, 10, the propionyl analogue of 3, showed excellent antiinflammatory activity and therefore represents a new lead structure for the development of injectable COX-2 specific inhibitors.

Novel 6,7-diphenyl-2,3,8,8a-tetrahydro-1H-indolizin-5-one analogues as cytotoxic agents.

A series of 6,7-diphenyl-2,3,8,8a-tetrahydro-1H-indolizin-5-one analogues were synthesized and evaluated for cytotoxic activity against eight human cancer cell lines. Compounds 18, 21, 28, 29, 30 and 31 showed cytotoxic activity with GI(50) values in the range of 2.1-8.1 microM concentration. Among these, compounds 21 and 28 exhibited good pharmacokinetic properties. These compounds were further evaluated for their in vivo efficacy in modified hollow fibre assay (HFA).

Novel indolo[2,1-b]quinazoline analogues as cytostatic agents: synthesis, biological evaluation and structure-activity relationship.

In our endeavor to design and synthesize novel anticancer agents, a new series of indoloquinazoline compounds were prepared and tested initially for anticancer activity in vitro against a panel of human cancer cell lines. Most of these compounds exhibited cytotoxic activity in in vitro screens. Compounds were selected and further evaluated using a modified Hollow Fiber Assay for their preliminary in vivo activity against 12 cell lines implanted in the subcutaneous and intraperitoneal compartments in mice. The results indicate that these compounds may constitute a new class of anticancer agents.

PMT13, a pyrimidone analogue of thiazolidinedione improves insulin resistance-associated disorders in animal models of type 2 diabetes.

AIM: To evaluate the antidiabetic and hypolipidaemic potential of a novel thiazolidinedione, PMT13, in different animal models of insulin resistance. METHODS: PPAR transactivation study was performed in HEK293T cells using ligand binding domains of PPARalpha, gamma and delta. Insulin-resistant db/db and ob/ob mice were treated orally with different doses of PMT13 at 0.3-10 mg/kg/day for 15 and 14 days respectively. Zucker fa/fa rats were treated with 3 mg/kg (p.o.) dose of the compound. Plasma glucose, triglyceride, free fatty acid and insulin levels were measured. Liver glucose 6-phosphatase (G6-Ptase) and adipose lipoprotein lipase activity was measured in treated mice. Isolated rat aortic preparations preconstricted with phenylephrine were used to study the vascular relaxation potential of PMT13 in presence of insulin. A 28-day oral toxicity study was performed in Wistar rats. RESULTS: PMT13 showed similar PPARgamma activation as rosiglitazone, but failed to show any activity against PPARalpha or PPARdelta. In obese and diabetic db/db and ob/ob mice, PMT13 showed better reduction in plasma glucose, triglyceride and insulin levels than rosiglitazone and an improvement in glucose tolerance. In insulin-resistant Zucker fa/fa rat model, PMT13 treatment showed better reduction in plasma triglyceride, free fatty acid and insulin levels than that of rosiglitazone. Treated mice showed decreased G6-Ptase activity in liver. The LPL activity was increased in post-heparin plasma and epididymal fat of treated db/db mice. In an isolated, precontracted rat aortic preparation, PMT13 treatment significantly increased insulin-induced relaxation. A 28-day oral toxicity study in rats showed no treatment-related adverse effects. CONCLUSION: Our studies indicate that PMT13 is a potent activator of PPARgamma with antidiabetic, hypolipidaemic and insulin-sensitizing properties. Additionally, PMT13 inhibited liver G6-Ptase activity and increased lipoprotein lipase activity. It showed improvement in insulin-induced vasorelaxation. The compound also showed a good safety margin. Therefore, PMT13 can be a potential drug candidate for future development.

Lack of effect of sucralfate on the absorption and pharmacokinetics of rosiglitazone.

The aim of the present study was to investigate the effect of sucralfate pretreatment on the pharmacokinetics of rosiglitazone following a single oral dose in healthy male volunteers. After an over night fast, and according to a randomized schedule, each volunteer (n = 9) received a single oral dose of rosiglitazone 8 mg (Avandia tablets, 4 mg x 2) with or without pretreatment of sucralfate 2 g (Recolfate tablets, 1 g x 2) in an open-label crossover study with a 2-week washout period. Plasma samples were collected over a period of 24 hours at regular intervals. Safety assessment included monitoring of the vital signs, blood parameters, and ECG. No statistically significant differences (p > 0.05) were observed for any of the calculated rosiglitazone pharmacokinetic parameters in the two treatment groups. The mean parameters, AUC0-infinity and Cmax, following rosiglitazone administration alone were 3825.02 ng x h/ml and 664.47 ng/ml, respectively, and for rosiglitazone administered after pretreatment with sucralfate were 4848.19 ng x h/ml and 624.88 ng/ml, respectively. The t(max) for rosiglitazone alone and for rosiglitazone after sucralfate treatments was 1.11 and 1.67 hours, respectively. The mean elimination half-life for rosiglitazone and rosiglitazone after sucralfate treatment was 4.35 and 4.51 hours, respectively. Fraction of rosiglitazone absorbed was calculated by the Wagner-Nelson method, and no statistically significant difference (p > 0.05) was observed for the two treatments. Since sucralfate pretreatment did not show any significant difference in the pharmacokinetics of rosiglitazone, no dose adjustment is warranted for rosiglitazone when it is administered with sucralfate.

(-)3-[4-[2-(Phenoxazin-10-yl)ethoxy]phenyl]-2-ethoxypropanoic acid [(-)DRF 2725]: a dual PPAR agonist with potent antihyperglycemic and lipid modulating activity.

(-)DRF 2725 (6) is a phenoxazine analogue of phenyl propanoic acid. Compound 6 showed interesting dual activation of PPAR alpha and PPAR gamma. In insulin resistant db/db mice, 6 showed better reduction of plasma glucose and triglyceride levels as compared to rosiglitazone. Compound 6 has also shown good oral bioavailability and impressive pharmacokinetic characteristics. Our study indicates that 6 has great potential as a drug for diabetes and dyslipidemia.

Novel antidiabetic and hypolipidemic agents. 5. Hydroxyl versus benzyloxy containing chroman derivatives.

Several thiazolidinediones having chroman moieties were synthesized and evaluated for their euglycemic and hypolipidemic activities. Some of the analogues having an aminoalkyl group as a linker between the chroman ring and 4-[5-(2,4-dioxo-1, 3-thiazolidinyl)methyl]phenoxy moiety seem to be better than troglitazone. In vitro transactivation assays of PPARgamma have been carried out with these glitazones to understand their molecular mechanism. For the first time we have found that some of the unsaturated thiazolidinediones are superior to their saturated counterpart in the in vivo assay. A more potent thiazolidinedione analogue than troglitazone is reported. Pharmacokinetic studies have shown that protection of the OH group in the chroman moiety leads to a decrease in metabolism, thereby resulting in a superior pharmacological profile.

High-performance liquid chromatographic determination of the insulin sensitizing agent DRF-2189 in rat plasma.

A high-performance liquid chromatographic method for the determination of DRF-2189, using troglitazone as internal standard, is described. A dichloromethane-ethyl acetate solvent mixture (6:4, v/v) was used as the extraction solvent. A Kromasil C18 column with a mobile phase consisting of 0.05 M phosphate buffer-acetonitrile-methanol (22.5:37.5:40) (pH 5.0) was used at a flow-rate of 1.0 ml/min. The eluate was monitored by using fluorescence detection with excitation and emission wavelengths at 292 nm and 325 nm, respectively. Ratio of peak area of analyte to internal standard was used for quantification of plasma samples. Using this method, the absolute recovery of DRF-2189 from rat plasma was >95% and the limit of quantitation was 50 ng/ml. The intra-day relative standard deviation (R.S.D.) ranged from 1.74 to 7.24% at 1 microg/ml and 1.86 to 3.83% at 10 microg/ml. The inter-day R.S.D.s were 8.34 and 4.91% at 1 and 10 microg/ml, respectively. The method was applied to measure plasma concentrations of DRF-2189 in pharmacokinetic studies in Wistar rats.

Probenecid affects the pharmacokinetics of ofloxacin in healthy volunteers.

OBJECTIVE: The aim of this randomised, double-blind, crossover study was to evaluate the effect of single-dose probenecid on the pharmacokinetics of ofloxacin in eight healthy male volunteers. METHODS: After an overnight fast, and according to a randomised sequence, each volunteer received either single oral ofloxacin 200mg (Hoechst Marion Roussel Ltd., Mumbai, India) or both ofloxacin (1 x 200mg) and probenecid (1 x 500mg) [Geno Pharmaceutical Ltd., Mumbai, India]. Blood samples were collected at regular intervals until 24 hours. Serum concentration versus time profiles for ofloxacin were generated and pharmacokinetic parameters were calculated by noncompartmental model analysis. RESULTS: Elimination half-life, mean residence time and area under the curve were significantly increased (4.86 vs 5.26h; 7.23 vs 7.95h; 10.28 vs 11.9 mg/L . h) [p < 0.01], whereas the total clearance was decreased (19.66 vs 16.95 L/h) [p < 0.01] in the presence of probenecid. Other pharmacokinetic parameters were not significantly affected by coadministration of probenecid. CONCLUSION: Concomitant administration of probenecid with ofloxacin may result in a decreased elimination half-life and consequently increased bioavailability of ofloxacin. Probenecid may be co-prescribed with ofloxacin; patients taking this combination should be closely monitored and dosage reduction should be considered if warranted in high-risk patients.

Disposition of rifampicin following intranasal and oral administration.

Leprosy is transmitted by dissemination of M.leprae which are lodged in the nose of the patients suffering from multibacillary (MB) type of the disease. Rifampicin, a potent bactericidal antileprotic drug is given orally to the patients with a view to make the infective cases non-infective. Earlier work by us has shown that intranasal administration of rifampicin helps in reducing the M.leprae load in the nose much faster than after conventional oral administration. In the present study, rifampicin concentrations in plasma/urine/nasal wash of healthy volunteers following oral and intranasal administration were determined. Following intranasal administration, rifampicin was not detectable in plasma and high concentrations were measured in the nasal wash. Following oral administration, rifampicin was not detectable in the nasal wash indicating that enough antibiotic levels are not available for clearing M.leprae from nose.

Tissue distribution and deposition of clofazimine in rat following subchronic treatment with or without rifampicin.

Tissue distribution and deposition characteristics of clofazimine (CAS 2030-63-9), an antileprotic drug in rats have been investigated following controlled sub-chronic administration (p.o.) for a period of 1-2 months. The drug was administered alone at a dose of 20 mg/kg body weight and in combination with rifampicin (CAS 13292-46-1) (20 mg/kg p.o.). Various tissues (liver, lung, spleen, small intestine, brain, heart, kidney, skin, stomach and subcutaneous fat) were analyzed for clofazimine in all the treated groups. High levels (range 0.9-3.6 mg/g of wet tissue) were observed in tissues having reticuloendothelial components. In other tissues the levels were relatively lower (range 3-114 micrograms/g of wet tissue). Histopathological studies revealed that clofazimine is deposited in many tissues in the form of reddish-orange crystals. Concomitant treatment with rifampicin did not significantly alter tissue distribution or deposition profile of clofazimine nor did it influence the histopathology.

In vitro protein binding and tissue distribution of D(+) usnic acid.

In vitro protein binding of D(+) usnic acid in rabbit plasma and purified bovine serum albumin was investigated by equilibrium dialysis. The drug was highly protein bound, approximately 99.2%, and the extent of protein binding remained constant at usnic acid concentrations in the range of 1-5 micrograms/ml. The extent of binding, however, tended to decrease at low albumin concentrations and higher drug concentrations; Scatchard plot analysis indicated the existence of two classes of binding sites with association constants of 34.3 x 10(-6) and 1.43 x 10(-6) M respectively. Tissue distribution studies of usnic acid were undertaken in rats following i.p. administration. Usnic acid was well distributed into well perfused organs. The tissue/plasma ratio in lungs was high, which could be advantageous in a therapeutic agent for pulmonary tuberculosis.